Preliminary Study Of Aging Biology In Daphnia Pulex

Senescence is a universal phenomenon,the ontogenyfrom fertilized eggs toold age.It is a spontaneous and inevitable process of structural and functional decline over time,which is often accompanied by the decrease of cell division and differentiation,the aggravation of oxidative damage of tissues and organs,the decrease of antioxidant capacity and the decline of reproductive ability.Daphnia pulex is an order of global,primarily,small freshwater crustaceans,which is commonly known as water fleawith two kinds of methods of reproduction in the life cycle of asexual reproduction(parthenogenesis)and sexual reproduction(sexual reproduction).This kind of small-bodied organism is a very important component of the zooplankton and is fundamental to the function,structure and biological productivity of natural water ecosystems.In this study,we use Daphnia pulex as the test organism,to investigate the aging biology of Daphnia pulex in morphology,enzymology,molecular biology and ultrastructure.In perspective of growth,before 20 days,the body length of individual increases with the increase of the days.When it reaches 20 days,individual stops growing,and even shrunk after 25 days.In perspective of breeding,reproduction is in increasing before 20 days and decreases after.Individual mortality appears in the 20 th day and speeds up after the 25 th day.Using scanning electron microscopy(SEM)and transmission electron microscopy(TEM)technology,when we obsever the ultrastructure of external form and gonads internal feature,we found that there is no significant difference in male Daphnia pulex but an obvious shinking in back of female in group 28 d.This structure may be caused by plasticity change of reproduction reducing.It may also be the reason to lead to shorten the length.Under the TEM,the aging leads to fewer lipids in the gonads and vacuoles.Swelling mitochondria absenced mitochondria crest and vesicles.It suggests that group 21 d has a significant characteristic of aging.However,the gonads age significantly faster than the external form.By enzymatic analysis,as individual age,ACP,antioxidase and antioxidant abilities show a downward trend.However,AKP,LPO,MDA presents a upward trend.It suggests that antioxidase activities gradually reduced,lead to the body of ROS and peroxide accumulate in individual aging.According to transcriptome results,we used RACE to clone the full-length cDNA of DpChit 1723 bp.Open reading frame(ROF)is 1371 bp and encoding 456 amino acides.By comparing homologous with the system of the evolutionary tree,we found that the closest kinship between crustaceans and insects.By real-time fluorescent quantitative PCR technology,we researched DpChit gene expression in different days and found that DpChit gene showed a trend of increased after decrease first,and the peak at the 20 th day,which showed a significant difference with the group 25d(P<0.05).In addition,using a variety of reported primers to explore HSP70 and HSP90,a variety of antioxidant enzyme gene(CAT,Mn-SOD,GSH-Px and GST)and Doublesex gene at transcription level by qPCR.It suggests that with individual aging,HSP70 and HSP90 show a decrease trend,and antioxidase gene expression is close to the antioxidant enzyme activities.However,Dsx gene expression offers upgrade firstly than descending latter tendency,and it is the highest point in the 15 th day.It indicates that after 15 days,the possiblity of transformation of reproductive is rising in Daphnia pulex.Proteomics is a term used to describe a dynamic gene regulation,for quantitative determination of gene expression levels of protein,and explain the mechanism of gene expression regulation.Results of Daphnia pulex aging proteomics analysis shows that,in the biological process,group 15 d mainly presents mitotic related protein expression,indicated the cell division is strong,however,group 20 d and 25 d,mainly show the peroxidation,suggests that at this time the body suffered a heavier peroxidation.From the perspective of the cellular component,group 15 d mainly shows some cytoplasmic related protein,but group 20 d and 25 d are lipid metabolism related protein expression.From the perspective of the molecular function,group 15 d performance to the ribosome structure and components related protein synthesis,nevertheless,after 20 days,indicidual mainly embodied in the oxidation and oxidordeuctase molecules.This suggest that group 15 d has a vigrous growth and rapid metabolism.After 20 days,individual presents a more obvious oxidation progress,and lipid metabolism is significantly associated with individual aging.